Under hypocalcemic conditions a unique substance is elevated from the in situ perfused rat liver which, when partially purified, non-competitively decreases adenylate cyclase activation by parathyroid hormone, epinephrine and glucagon in several tissues and in several species; higher concentrations of this substance decreased basal, and fluoride activation of the enzyme also. The inhibitor is a small molecular weight acid stable and alkali labile substance. In the proposed study an attempt will be made to isolate, purify, and chemically characterize this substance and to study its distribution in the cell and in various tissues and body fluids under a variety of physiological conditions. Once purified, we intend to produce antibodies against this compound and develop a radioimmunoassay for it. The role of this inhibitor in pathological conditions related to cyclic nucleotide-mediated hormone responsiveness in hypocalcemic states will be investigated. Utilizing infusion techniques in rats, additional studies will be done to determine the effects of the inhibitor in vitro on a number of enzymes that have been shown to be involved either in the breakdown of cyclic nucleotides, or whose activity is altered by cyclic AMP or cyclic GMP. These include phosphodiesterases, protein kinase, guanylate cyclase, phosphorylase, glycogen synthase, phosphatase and ATPase. The levels of cyclic AMP and the inhibitor will be measured in the perfused rat liver preparations in response to hormones and exogenously administered cyclic nucleotides under a variety of conditions. BIBLIOGRAPHIC REFERENCES: Lo, H.K., Lehotay, D.C., Katz, D. and Levey, G.S.: Parathyroid hormone mediated incorporation of 32P-orthophosphate into phosphatidic acid and phosphatidylinositol in renal cortical slices. Endocrine Research Communications 3:377-385 (1976). Lo, H., Lehotay, D.C. and Levey, G.S.: Parathyroid hormone-mediated incorporation of 32P-orthophosphate into phosphotidic acid and phosphotidylinositol in renal cortical slices. Clinical Res. 25:32A, 1977.